Our laboratory is committed to the devising ways to prevent, diagnose and treat implant-associated orthopaedic infections with a focus on those caused by Staphylococcus aureus. While S. aureus causes only 30% of these infections it causes the most challenging implant-associated infections generally requiring removal of the implant and extensive antibiotic therapy. Following expensive and prolonged interventions, the success rate of therapy is only about 50%, leaving thousands of patients with limited mobility each year. Considering the gravity of these infections, an area that has lagged is diagnostics which is based on 24- 48 hour cultures. The advent of powerful molecular techniques like PCR and MALDI-TOF MS has helped speed up the identification of infecting bacteria but these methods still require that the pathogen be present in the sample taken for analysis. Sample-taking can be a considerable issue for deep-seated infections, often requiring expensive image-guided biopsies or surgery of the infected site. Here we propose to develop a new method for diagnosis of implant-associated S. aureus infections that obviates the need for obtaining a pathogen-containing sample and requires only a small sample of whole blood. The proposed method also introduces a new analytical fluid that eliminates many of the cross-reactivity and interference-causing factors present in serum or plasma. Harvested peripheral blood mononuclear cells are cultured in defined cell culture medium where a small subpopulation of recently activated B-cells, circulating plasmablasts, express their antibodies. We then identify the secreted antibodies by their activity in immunoassays specific for S. aureus proteins. This approach to bacterial diagnostics also solves the problem that plagues most antibody-based diagnostic tests for micro-organisms, the presence of serum-borne antibodies from prior exposure to the pathogen. This approach has been used with some success for viral infections, but almost no work has been published for bacteria. The fundamental problem has been how to identify those antigens that are certain to elicit antibody production by the host immune response in the context of heterogeneity in both the pathogen and the host. We propose to test our method using a population of patients with suspected implant-associated infections to simultaneously prove the utility of our approach in a clinical setting and optimize the antigen repertoire to maximize sensitivity and specificity. With a firm foundation built on proven clinical performance and assay simplification we can then address other indications for S. aureus like sepsis, pneumonia and atopic dermatitis, and ultimately other infections caused by other pathogens. The research-friendly format we use in the proposed experiments can also be readily simplified to make the process rapid and inexpensive to meet the clinical need.